Although the mTOR-4E-BP1 signaling pathway is implicated in aging and aging-related disorders, the role of 4E-BP1 in regulating human stem cell homeostasis remains largely unknown. Here, we report that the expression of 4E-BP1 decreases along with the senescence of human mesenchymal stem cells (hMSCs). Genetic inactivation of 4E-BP1 in hMSCs compromises mitochondrial respiration, increases mitochondrial reactive oxygen species (ROS) production, and accelerates cellular senescence. Mechanistically, the absence of 4E-BP1 destabilizes proteins in mitochondrial respiration complexes, especially several key subunits of complex III including UQCRC2. Ectopic expression of 4E-BP1 attenuates mitochondrial abnormalities and alleviates cellular senescence in 4E-BP1-deficient hMSCs as well as in physiologically aged hMSCs. These f indings together demonstrate that 4E-BP1 functions as a geroprotector to mitigate human stem cell senescence and maintain mitochondrial homeostasis, particularly for the mitochondrial respiration complex III, thus providing a new potential target to counteract human stem cell senescence.
Mesenchymal Stem Cells/physiology* ; Cellular Senescence ; Homeostasis ; Cell Cycle Proteins/metabolism* ; Adaptor Proteins, Signal Transducing/metabolism* ; Mitochondria/metabolism* ; Electron Transport Complex III/metabolism* ; Humans ; Cells, Cultured

Cell cycle plays a crucial role in cell development. Cell cycle progression is mainly regulated by cyclin dependent kinase (CDK), cyclin and endogenous CDK inhibitor (CKI). Among these, CDK is the main cell cycle regulator, binding to cyclin to form the cyclin-CDK complex, which phosphorylates hundreds of substrates and regulates interphase and mitotic progression. Abnormal activity of various cell cycle proteins can cause uncontrolled proliferation of cancer cells, which leads to cancer development. Therefore, understanding the changes in CDK activity, cyclin-CDK assembly and the role of CDK inhibitors will help to understand the underlying regulatory processes in cell cycle progression, as well as provide a basis for the treatment of cancer and disease and the development of CDK inhibitor-based therapeutic agents. This review focuses on the key events of CDK activation or inactivation, and summarizes the regulatory processes of cyclin-CDK at specific times and locations, as well as the progress of research on relevant CDK inhibitor therapeutics in cancer and disease. The review concludes with a brief description of the current challenges of the cell cycle process, with the aim to provide scientific references and new ideas for further research on cell cycle process.
Cyclin-Dependent Kinases/metabolism* ; Cyclins/metabolism* ; Protein Serine-Threonine Kinases ; Cell Cycle Proteins/metabolism* ; Cell Cycle/physiology* ; Cyclin-Dependent Kinase 2

Proteasome inhibitors have shown remarkable success in the treatment of hematologic neoplasm. There has been a lot of attention to applying these drugs for solid tumor treatment. Recent preclinical study has signified the effectiveness on cell proliferation inhibition in lung adenocarcinoma treated by carfilzomib (CFZ), a second generation proteasome inhibitor. However, no insight has been gained regarding the mechanism. In this study, we have systematically investigated the CFZ functions in cell proliferation and growth, cell cycle arrest, and apoptosis in lung adenocarcinoma cells. Flow cytometry experiments showed that CFZ significantly induced G2/M cell cycle arrest and apoptosis in lung adenocarcinoma. MTS and colony formation assays revealed that CFZ substantially inhibited survival of lung adenocarcinoma cells. All results were consistently correlated to the upregulation expression of Gadd45a, which is an important gene in modulating cell cycle arrest and apoptosis in response to physiologic and environmental stresses. Here, upregulation of Gadd45a expression was observed after CFZ treatment. Knocking down Gadd45a expression suppressed G2/M arrest and apoptosis in CFZ-treated cells, and reduced cytotoxicity of this drug. The protein expression analysis has further identified that the AKT/FOXO3a pathway is involved in Gadd45a upregulation after CFZ treatment. These findings unveil a novel mechanism of proteasome inhibitor in anti-solid tumor activity, and shed light on novel preferable therapeutic strategy for lung adenocarcinoma. We believe that Gadd45a expression can be a highly promising candidate predictor in evaluating the efficacy of proteasome inhibitors in solid tumor therapy.
Adenocarcinoma of Lung/pathology* ; Apoptosis/drug effects* ; Cell Cycle Checkpoints/drug effects* ; Cell Cycle Proteins/genetics* ; Cell Line, Tumor ; Forkhead Box Protein O3/physiology* ; Gene Expression Regulation, Neoplastic/drug effects* ; Humans ; Lung Neoplasms/pathology* ; Oligopeptides/pharmacology* ; Proto-Oncogene Proteins c-akt/physiology* ; Up-Regulation

BACKGROUND: The clinical trials emerged centromere protein E inhibitor GSK923295 as a promising anticancer drug, but its function in hepatocellular carcinoma (HCC) remain needs to be fully elucidated, especially as chemotherapy after hepatectomy for liver tumors. We aimed to describe anti-HCC activities of GSK923295 and compare its antiproliferative effects on liver regeneration after partial hepatectomy (PH). METHODS: All subjects were randomized to treatment with either vehicle or GSK923295. Antitumor activity of GSK923295 was assessed by xenograft growth assays. The C57BL/6 mice were subjected to 70% PH and the proliferation was calculated by liver coefficient, further confirmed by immunohistochemistry. The proliferation and cell cycle analysis of liver cell AML12 and HCC cells LM3, HUH7, and HepG2 were investigated using the cell counting kit-8 assay and Flow Cytometry. The chromosome misalignment and segregation in AML12 cells were visualized by immunofluorescence. RESULTS: Treatment with GSK923295 induced antiproliferation in HCC cell lines. It also caused delay on HCC tumor growth instead of regression both in a HCC cell line xenograft model and patient-derived tumor xenograft model. With microarray analysis, CENtromere Protein E was gradually increased in mouse liver after PH. Exposure of liver cells to GSK923295 resulted in delay on a cell cycle in mitosis with a phenotype of misaligned chromosomes and chromosomes clustered. In 70% PH mouse model, GSK923295 treatment also remarkably reduced liver regeneration in later stage, in parallel with the mitotic marker phospho-histone H3 elevation. CONCLUSION The anticancer drug GSK923295 causes a significant delay on HCC tumor growth and liver regeneration after PH in later stage.
Animals ; Antineoplastic Agents ; therapeutic use ; Blotting, Western ; Bridged Bicyclo Compounds, Heterocyclic ; therapeutic use ; Carcinoma, Hepatocellular ; drug therapy ; surgery ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Chromosomal Proteins, Non-Histone ; antagonists & inhibitors ; Electrophoresis, Polyacrylamide Gel ; Female ; Fluorescent Antibody Technique ; Humans ; Immunohistochemistry ; Liver Neoplasms ; drug therapy ; surgery ; Liver Regeneration ; physiology ; Mice ; Mice, Inbred C57BL ; Real-Time Polymerase Chain Reaction ; Sarcosine ; analogs & derivatives ; therapeutic use ; Xenograft Model Antitumor Assays

BACKGROUNDBrain acid soluble protein 1 (BASP1) is identified as a novel potential tumor suppressor in several cancers. However, its role in thyroid cancer has not been investigated yet. In the present study, the antitumor activities of BASP1 against the growth and migration of thyroid cancer cells were evaluated.

METHODSBASP1 expression in thyroid cancer tissues and normal tissues were examined by immunohistochemical staining and the association between its expression and prognosis was analyzed. pcDNA-BASP1 carrying full length of BASP1 cDNA was constructed to restore the expression of BASP1 in thyroid cancer cell lines (BHT-101 and KMH-2). The cell proliferation in vitro and in vivo was evaluated by WST-1 assay and xenograft tumor models, respectively. Cell cycle distribution after transfection was analyzed using flow cytometry. Cell apoptosis after transfection was examined by annexin V/propidium iodide assay. The migration was examined using transwell assay.

RESULTSBASP1 expression was abundant in normal tissues while it is significantly decreased in cancer tissues (P = 0.000). pcDNA-BASP1 restored the expression of BASP1 and significantly inhibited the growth of BHT-101 and KMH-2 cells as well as xenograft tumors in nude mice (P = 0.000). pcDNA-BASP1 induced G1 arrest and apoptosis in BHT-101 and KMH-2 cells. In addition, pcDNA-BASP1 significantly inhibited the cell migration.

CONCLUSIONSDownregulation of BASP1 expression may play a role in the tumorigenesis of thyroid cancer. Restoration of BASP1 expression exerted extensive antitumor activities against growth and migration of thyroid cancer cells, which suggested that BASP1 gene might act as a potential therapeutic agent for the treatment of thyroid cancer.

Aged ; Animals ; Apoptosis ; genetics ; physiology ; Calmodulin-Binding Proteins ; genetics ; metabolism ; Cell Cycle ; genetics ; physiology ; Cell Line, Tumor ; Cell Movement ; genetics ; physiology ; Cell Proliferation ; genetics ; physiology ; Cytoskeletal Proteins ; genetics ; metabolism ; Female ; Gene Expression Regulation, Neoplastic ; genetics ; physiology ; Humans ; Male ; Membrane Proteins ; genetics ; metabolism ; Mice ; Mice, Nude ; Middle Aged ; Nerve Tissue Proteins ; genetics ; metabolism ; Repressor Proteins ; genetics ; metabolism ; Thyroid Neoplasms ; metabolism ; pathology ; Xenograft Model Antitumor Assays

BACKGROUNDRetinal degenerative diseases are the leading causes of blindness in developed world. Retinal progenitor cells (RPCs) play a key role in retina restoration. Triamcinolone acetonide (TA) is widely used for the treatment of retinal degenerative diseases. In this study, we investigated the role of TA on RPCs in hypoxia condition.

METHODSRPCs were primary cultured and identified by immunofluorescence staining. Cells were cultured under normoxia, hypoxia 6 h, and hypoxia 6 h with TA treatment conditions. For the TA treatment groups, after being cultured under hypoxia condition for 6 h, RPCs were treated with different concentrations of TA for 48-72 h. Cell viability was measured by cell counting kit-8 (CCK-8) assay. Cell cycle was detected by flow cytometry. Western blotting was employed to examine the expression of cyclin D1, Akt, p-Akt, nuclear factor (NF)-κB p65, and caspase-3.

RESULTSCCK-8 assays indicated that the viability of RPCs treated with 0.01 mg/ml TA in hypoxia group was improved after 48 h, comparing with control group (P < 0.05). After 72 h, the cell viability was enhanced in both 0.01 mg/ml and 0.02 mg/ml TA groups compared with control group (all P < 0.05). Flow cytometry revealed that there were more cells in S-phase in hypoxia 6 h group than in normoxia control group (P < 0.05). RPCs in S and G2/M phases decreased in groups given TA, comparing with other groups (all P < 0.05). There was no significant difference in the total Akt protein expression among different groups, whereas upregulation of p-Akt and NF-κB p65 protein expression and downregulation of caspase-3 and cyclin D1 protein expression were observed in 0.01 mg/ml TA group, comparing with hypoxia 6 h group and control group (all P < 0.05).

CONCLUSIONLow-dose TA has anti-apoptosis effect on RPCs while it has no stimulatory effect on cell proliferation.

Animals ; Apoptosis ; drug effects ; physiology ; Caspase 3 ; metabolism ; Cell Cycle ; drug effects ; physiology ; Cell Hypoxia ; drug effects ; physiology ; Cell Proliferation ; drug effects ; physiology ; Cell Survival ; drug effects ; physiology ; Cells, Cultured ; Cyclin D1 ; metabolism ; NF-kappa B ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Rats ; Rats, Sprague-Dawley ; Retina ; cytology ; Stem Cells ; cytology ; drug effects ; Triamcinolone Acetonide ; pharmacology

The ubiquitin-proteasome system (UPS) is a proteasome system widely present in the human body, which is composed of ubiquitin (Ub), ubiquitin activating enzymes (E1), ubiquitin conjugating enzymes (E2), ubiquitin protein ligases (E3), 26S proteasome, and deubiquitinating enzymes (DUBs) and involved in cell cycle regulation, immune response, signal transduction, DNA repair as well as protein degradation. Sperm DNA is vulnerable to interference or damage in the progression of chromosome association and homologous recombination. Recent studies show that UPS participates in DNA repair in spermatogenesis by modulating DNA repair enzymes via ubiquitination, assisting in the identification of DNA damage sites, raising damage repair-related proteins, initiating the DNA repair pathway, maintaining chromosome stability, and ensuring the normal process of spermatogenesis.
Cell Cycle Proteins ; physiology ; DNA Damage ; DNA Repair ; physiology ; Humans ; Male ; Proteasome Endopeptidase Complex ; physiology ; Signal Transduction ; physiology ; Spermatogenesis ; physiology ; Spermatozoa ; Ubiquitin ; physiology ; Ubiquitin-Conjugating Enzymes ; physiology ; Ubiquitin-Protein Ligases ; physiology ; Ubiquitination

BACKGROUNDThe Notch-regulated ankyrin repeat protein (NRARP) is recently found to promote proliferation of breast cancer cells. The role of NRARP in carcinogenesis deserves extensive investigations. This study attempted to investigate the expression of NRARP in thyroid cancer tissues and assess the influence of NRARP on cell proliferation, apoptosis, cell cycle, and invasion in thyroid cancer.

METHODSThirty-four cases with thyroid cancer were collected from the Department of General Surgery, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine between 2011 and 2012. Immunohistochemistry was used to detect the level of NRARP in cancer tissues. Lentivirus carrying NRARP-shRNA (Lenti-NRARP-shRNA) was applied to down-regulate NRARP expression. Cell viability was tested after treatment with Lenti-NRARP-shRNA using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Apoptosis and cell cycle distribution were determined by flow cytometry. Cell invasion was tested using Transwell invasion assay. In addition, expressions of several cell cycle-associated and apoptosis-associated proteins were examined using Western blotting after transfection. Student's t-test, one-way analysis of variance (ANOVA), or Kaplan-Meier were used to analyze the differences between two group or three groups.

RESULTSNRARP was highly expressed in thyroid cancer tissues. Lenti-NRARP-shRNA showed significantly inhibitory activities against cell growth at a multiplicity of infection of 10 or higher (P < 0.05). Lenti-NRARP-shRNA-induced G1 arrest (BHT101: 72.57% ± 5.32%; 8305C: 75.45% ± 5.26%) by promoting p21 expression, induced apoptosis by promoting bax expression and suppressing bcl-2 expression, and inhibited cell invasion by suppressing matrix metalloproteinase-9 expression.

CONCLUSIONDownregulation of NRARP expression exerts significant antitumor activities against cell growth and invasion of thyroid cancer, that suggests a potential role of NRARP in thyroid cancer targeted therapy.

Adult ; Aged ; Animals ; Apoptosis ; genetics ; physiology ; Cell Cycle ; genetics ; physiology ; Cell Line, Tumor ; Cell Proliferation ; genetics ; physiology ; Cell Survival ; genetics ; physiology ; Female ; Humans ; In Vitro Techniques ; Kaplan-Meier Estimate ; Male ; Mice ; Mice, Nude ; Middle Aged ; Neoplasm Proteins ; genetics ; metabolism ; Proteins ; genetics ; metabolism ; RNA, Small Interfering ; genetics ; Thyroid Neoplasms ; genetics ; metabolism ; mortality ; pathology

To explore the role of the special AT rich sequence binding protein-1 (SATB1) and ribonucleotide reductase M2 (RRM2) in enhancing malignant progression of non-Hodgkin lymphoma (NHL). 
 Methods: A total of 42 NHL and 42 chronic lymphadenitis patients were recruited. The protein expressions of SATB1 and RRM2 in cervical lymph nodes were determined by Western blot. After overexpression of SATB1, siSATB1 or siRRM2, the mRNA levels of SATB1 and RRM2 in cells were analyzed via RT-PCR, the cell proliferation was evaluated via MTT and EdU assays, while the migration and invasion of cells were assessed by transwell assays.
 Results: Compared with chronic lymphadenitis, the expressions of SATB1 and RRM2 in NHL patients were up-regulated. There was positive correlation between SATB1 and RRM2 in NHL patients. RRM2 mRNA level was up-regulated after transfection of SATB1 and down-regulated after transfection of siSATB1. Overexpression of SATB1 increased tumor cell proliferation, migration and invasion, while knockdown of RRM2 reversed those phenomena.
 Conclusion: SATB1 functions as an oncogene and promotes tumor cell proliferation, migration and invasion by up-regulation of RRM2 in NHL.
Cell Cycle Proteins ; genetics ; physiology ; Cell Movement ; genetics ; Cell Proliferation ; genetics ; Down-Regulation ; Gene Expression Regulation, Neoplastic ; genetics ; Humans ; Lymph Nodes ; chemistry ; Lymphoma, Non-Hodgkin ; Matrix Attachment Region Binding Proteins ; genetics ; physiology ; Neoplasm Invasiveness ; genetics ; Oncogenes ; genetics ; physiology ; RNA, Messenger ; RNA, Small Interfering ; Ribonucleoside Diphosphate Reductase ; Ribonucleotide Reductases ; genetics ; physiology ; Signal Transduction ; Transcription Factors ; Transcriptional Activation ; Transfection ; Tumor Cells, Cultured ; Up-Regulation

To investigate effects of MARCH6 gene knockdown on MCF-7 cell proliferation and cell cycle.
 Methods: 293T cells were transfected with MARCH6 shRNA lentivirus. Fluorescence microscope was used to observe and verify the transfection efficiency. The initial effect of the MARCH6 gene knockdown in MCF-7 cells was observed via fluorescence microscope. Real-time PCR and Western blot were used to detect the expression of MARCH6. MTT and BrdU assay were used to examine cell proliferation, and staining flow cytometry was used to analyze cycle distribution of MCF-7 cells.
 Results: MARCH6 shRNA lentivirus was successfully transfected and about 80% of the cells expressed green fluorescent in comparison of the control. About 90% of the cells showed green fluorescence. The mRNA and protein in MCF-7 cells were transcription and expression of protein was significantly decreased after the transfection of MARCH6 shRNA lentivirus accompanied by a decrease in MCF-7 cell proliferation (P<0.01). Flow cytometry showed that the cell cycles were inhibited at the G1 phase and the proliferation index was significantly reduced.
 Conclusion: Knockdown of MARCH6 gene by RNA interference inhibits the proliferation of MCF-7 cells, suggesting that the expression of MARCH6 promotes proliferation of breast cancer cells through regulation of the cell cycle.
Adenocarcinoma ; genetics ; Breast Neoplasms ; genetics ; Cell Cycle ; Cell Division ; Cell Proliferation ; genetics ; Female ; G1 Phase ; genetics ; Gene Knockdown Techniques ; Humans ; Hyperplasia ; Lentivirus ; MCF-7 Cells ; physiology ; Membrane Proteins ; physiology ; RNA Interference ; RNA, Messenger ; RNA, Small Interfering ; Real-Time Polymerase Chain Reaction ; Transfection ; Ubiquitin-Protein Ligases ; physiology